Cold Inactivation of Phosphoenolpyruvate Carboxylase and Pyruvate Orthophosphate Dikinase from the C4 Perennial Plant Atriplex halimus

Phosphoenolpyruvate carboxylase (PEPC) and pyruvate orthophosphate dikinase (PPDK) cold inactivation was studied in leaf extracts from Atriplex halimus L. Both enzyme activities gradually reduced as the temperature and the total soluble protein decreased. Mg²⁺ at a concentration of 10 mM stabilized PEPC and PPDK activities against cold inactivation. At low Mg²⁺ concentration (4 mM), PEPC was strongly protected by phosphoenolpyruvate, glucose-6-phosphate, and, partially, byL-malate, while PPDK was protected by PEP, but not by its substrate, pyruvate. High concentrations of compatible solutes (glycerol, betaine, proline, sorbitol and trehalose) proved to be good protectants for both enzyme activities against cold inactivation. When illuminated leaves were exposed to low temperature, PPDK was partially inactivated, while the activity of PEPC was not altered.     

Growth and N2 fixation in mixed cropping of Medicago arborea and Atriplex halimus grown on a salt-affected soil using a 15N tracer technique

Abstract

The objective of this study was to evaluate dry matter (DM), nitrogen yield, N₂ fixation (Ndfa) and soil N uptake (Ndfs) in the shrubby medic (Medicago arborea) and saltbush (Atriplex halimus) grown in pots either solely or in a mixture on a salt-affected soil, using ¹⁵N dilution method. The combined DM of both species was considerably higher than that of solely grown shrubs. The inclusion of saltbush in the mixed cropping system decreased Ndfs by shrubby medic and enhanced % Ndfa without affecting amounts of N₂ fixed. It can be concluded that the use of mixed cropping system of shrubby medic and saltbush could be a promising bio-saline agricultural approach to utilize salt affected soils in terms of forage yield and N₂ fixation.     

An inland and a coastal population of the Mediterranean xero-halophyte species Atriplex halimus L. differ in their ability to accumulate proline and glycinebetaine in response to salinity and water stress

Soil salinity and drought compromise water uptake and lead toosmotic adjustment in xero-halophyte plant species. These importantenvironmental constraints may also have specific effects onplant physiology. Stress-induced accumulation of osmocompatiblesolutes was analysed in two Tunisian populations of the Mediteraneanshrub Atriplex halimus L.—plants originating from a salt-affectedcoastal site (Monastir) or from a non-saline semi-arid area(Sbikha)—were exposed to nutrient solution containingeither low (40 mM) or high (160 mM) doses of NaCl or 15% polyethyleneglycol. The low NaCl dose stimulated plant growth in both populations.Plants from Monastir were more resistant to high salinity andexhibited a greater ability to produce glycinebetaine in responseto salt stress. Conversely, plants from Sbikha were more resistantto water stress and displayed a higher rate of proline accumulation.Proline accumulated as early as 24 h after stress impositionand such accumulation was reversible. By contrast, glycinebetaineconcentration culminated after 10 d of stress and did not decreaseafter the stress relief. The highest salt resistance of Monastirplants was not due to a lower rate of Na⁺ absorption; plantsfrom this population exhibited a higher stomatal conductanceand a prodigal water-use strategy leading to lower water-useefficiency than plants from Sbikha. Exogenous application ofproline (1 mM) improved the level of drought resistance in Monastirplants through a decrease in oxidative stress quantified bythe malondialdehyde concentration, while the exogenous applicationof glycinebetaine improved the salinity resistance of Sbikhaplants through a positive effect on photosystem II efficiency. Key words: Atriplex halimus, glycinebetaine, halophyte, NaCl, osmotic adjustment, proline, salinity, water stress     

The moderately halophilic bacterium Halomonas maura is a free-living diazotroph

Halomonas maura is a moderately halophilic bacterium which lives in saline soils and synthesises an exopolysaccharide known as mauran. Strain S-31T grew in a nitrogen-free medium under an N2 atmosphere; the acetylene reduction assay proved positive under specific conditions. We identified the nifH gene in this strain by using degenerate oligonucleotides designed from highly preserved gene sequences obtained from the alignment of a large number of nifH sequences from different microorganisms. Our results lead us to conclude that H. maura is capable of fixing nitrogen under microaerobic conditions.     

安徽定远盐矿可培养嗜盐微生物多样性

【背景】嗜盐微生物多生活于高盐环境,具有独特的生理代谢特征,是一类重要的极端环境微生物资源。【目的】为更好地认识我国陆相盐矿的嗜盐微生物多样性组成,更好地开发利用嗜盐微生物资源积累丰富的微生物菌种。【方法】对安徽定远盐矿盐芯样品进行嗜盐微生物的纯培养分离,并对所分离菌株进行基于16SrRNA基因的测序和序列相似性分析,并对所分离菌株进行物种多样性分析。在此基础上,对代表菌株进行菌落形态和耐盐度及酶活测定。【结果】通过纯培养共分离获得了嗜盐微生物264株,其中嗜盐古菌150株,占56.8%;嗜盐细菌114株,占43.2%。嗜盐古菌物种分别来自于Halorubrum、 Halopenitus、 Haloterrigena、 Natrinema、 Natronoarchaeum和Natronomonas等6个属;嗜盐细菌物种分别来自于Pseudomonas、Aliifodinibius、Halobacillus、Halomonas和Halospina等5个属。通过代表菌株的酶活平板检测,发现产胞外蛋白酶菌株1株,酯酶1株,淀粉酶2株;能液化明胶菌株2株。在物种多样性组成方面,发现嗜盐古菌的物种多样性指数高于嗜盐细菌。【结论】本研究对我国安徽定远陆相盐矿的可培养嗜盐微生物多样性进行探究,积累了丰富的嗜盐微生物菌株资源。     

A novel moderately halophilic bacterium for decolorizing azo dye under high salt condition

Halomonas sp strain GTW was newly isolated from coastal sediments contaminated by chemical wastewater and was identified to be a member of the genus Halomonas by 16S rDNA sequence analysis and physical and biochemical tests. The optimal decolorization conditions were as follows: temperature 30°C, pH 6.5.0–8.5, NaCl 10–20% (w/v) and the optimal carbon source was yeast exact. The results of experiments demonstrated that the bacteria could decolorize different azo dyes under high salt concentration conditions, and the decolorization rate of five tested azo dyes could be above 90% in 24 h. The exploitation of the salt-tolerant bacteria in the bio-treatment system would be a great improvement of conventional biological treatment systems and the bio-treatment concept.     

Sampling Atriplex bladders: a comparison of methods

Abstract. A major objection to the convenient sampling of Chenopodiacean bladders by brushing in aqueous solutions is the suspected leakage of ions from the leaf blade. To overcome this problem, a new method of sampling bladder hair is described, which also achieves rapid and quantitative separation and, at the same time, yields bladder samples of undoubtedly high purity. Leakage is stopped by the aid of liquid nitrogen. Comparison of this method to removal of bladders by brushing in aquenous solutions effectively confirm the validity of the customary method beyond dispute.     

Purification and characterisation of two extremely halotolerant xylanases from a novel halophilic bacterium

The present work reports for the first time the purification and characterisation of two extremely halotolerant endo-xylanases from a novel halophilic bacterium, strain CL8. Purification of the two xylanases, Xyl 1 and 2, was achieved by anion exchange and hydrophobic interaction chromatography. The enzymes had relative molecular masses of 43 kDa and 62 kDa and pI of 5.0 and 3.4 respectively. Stimulation of activity by Ca²⁺, Mn²⁺, Mg²⁺, Ba²⁺, Li²⁺, NaN₃ and isopropanol was observed. The K_m and V_max values determined for Xyl 1 with 4-O-methyl-d-glucuronoxylan are 5 mg/ml and 125,000 nkat/mg respectively. The corresponding values for Xyl 2 were 1 mg/ml and 143,000 nkat/mg protein. Xylobiose and xylotriose were the major end products for both endoxylanases. The xylanases were stable at pH 4–11 showing pH optima around pH 6. Xyl 1 shows maximal activity at 60°C, Xyl 2 at 65°C (at 4 M NaCl). The xylanases showed high temperature stability with half-lives at 60°C of 97 min and 192 min respectively. Both xylanases showed optimal activity at 1 M NaCl, but substantial activity remained for both enzymes at 5 M NaCl.Communicated by W.D. Grant     

A bacteriophage of a moderately halophilic bacterium

A bacteriophage of an aerobic, gram-negative, rod-shaped halophilic bacterium, provisionally named Pseudomonas sp. G3, is described. The phage has a head and a tail and is similar in appearance to Salmonella phage Beccles. It infects its bacterial host at all salt concentrations in which the bacteirum is able to grow. In contrast to phages of halophilic archaebacteria, the newly-described phage is relatively stable in the absence of salt. It also infects Vibrio costicola and two unidentified halophilic eubacteria.Abbreviations PPT proteose peptone-tryptone medium - pfu plaque-forming unit - G+C guanine + cytidine content, mol %     

艾丁湖可培养嗜盐菌多样性及功能酶、抗菌活性筛选

【目的】探究艾丁湖可培养嗜盐菌的多样性、功能酶活性以及抗菌活性,进一步了解其次级代谢产物情况,为新型生物活性物质的发掘提供依据。【方法】选用以20种糖及糖的衍生物作为唯一碳源的寡营养培养基从艾丁湖5个样点中分离得到298株嗜盐菌,根据形态特征去重复后,选取62株菌运用16S r RNA基因系统发育分析的方法研究样品中嗜盐菌的多样性;从不同类群选取22株代表菌株,采用点接法进行3种功能酶的筛选,运用平板对峙法检测代表菌株对12种病原菌的抗菌活性。【结果】从5%、10%和15%3个盐浓度中分别分得221、54和23株嗜盐菌,获得的嗜盐菌分布在9个科18个属;其中放线菌分布于4个属,细菌分布于14个属;Nocardiopsis和Pontibacillus属为艾丁湖可培养嗜盐菌的优势类群,分别占17.7%和16.1%;有15株嗜盐菌相似性低于98.5%,可能为潜在新种。所选取的22株代表菌株中,分别有68.2%、22.7%和72.7%的实验菌株具有蛋白酶、淀粉酶和酯酶活性;45.5%的代表菌株对12种病原菌表现出了抗至少1种病原菌的活性,其中一株Nocardiopsis属放线菌能抗9种病原菌,表现出了广谱的抗菌活性。【结论】新疆艾丁湖土样中嗜盐菌的多样性较丰富,而且具有较好的生物活性,为后续进一步研究其次级代谢产物提供了依据。     

Purification and characterization of three proteins from a halophilic sulfate-reducing bacterium,Desulfovibrio salexigens

Summary Hydrogenase, desulfoviridin and molybdenum proteins have been isolated from a halophilic sulfate-reducing bacteria,Desulfovibrio salexigens strain British Guiana. At least 50% of the hydrogenase was found to be located in the periplasm. The hydrogenase has a typical absorption spectrum, a 400/280 nm ratio of 0.28, a molecular weight by sedimentation equilibrium of 81 000 and is composed of two subunits. It has one nickel, one selenium and 12 iron atoms per molecule. The sulfite reductase has a typical desulfoviridin absorption spectrum, a molecular weight of 191 000 and iron and zinc associated with it. The molybdenum-iron protein is gray-green in color and exhibits an absorbtion spectrum with peaks around 612, 410, 275 nm and a shoulder at 319 nm. It is composed of subunits of approximately 13 250 and has an approximate molecular weight of 110 000. Three molybdenum and 20 iron atoms are found associated with it.An extensive study of these three proteins will allow a better understanding of the function of these enzymes and also of their possible role in microbially caused corrosion.     

Isolation of an obligately chemolithoautotrophic,halophilic and aerobic hydrogen-oxidizing bacterium from marine environment

An obligately chemolithoautotrophic and aerobic hydrogen-oxidizing bacterium was isolated from seawater of the Shonan Coast, Kanagawa Pref., Japan. The isolate was a Gram-negative, comma-shaped rod cell measuring 0.2 to 0.5 by 1 to 2 m. The cells occurred singly and were motile by a polar flagellum. The deoxyribonucleic acid base composition of the isolate was 44.1 mol% guanine plus cytosine. The optimal temperature for autotrophic growth on H₂–O₂–CO₂ was around 37°C, and no growth was observed at 5° C or 45° C. The optimal pH for growth was around 6.5. NaCl was required for growth with an optimum of 0.5 M. Elemental sulfur, thiosulfate or tetrathionate was utilized, as well as molecular hydrogen, as the sole energy source. No heterotrophic growth was observed on organic media tested. To our knowledge, this is the first report of the isolation of a marine, aerobic hydrogen-oxidizing bacterium, and of an obligately chemolithoautotrophic, mesophilic and aerobic hydrogen-oxidizing bacterium.     

The effect of fruit age on seed germinability of a heterocarpic species, Atriplex sagittata

Atriplex sagittata is an annual heterocarpic plant that produces three different fruit types (termed A, B, and C). To examine the consequence of heterocarpy on germinability patterns over long time periods, we compared seed germinability of different fruit types that had been stored for up to 8 years. While germinability of non-dormant type C fruits in distilled water was high (up to 100 %) in the first 2 years, it rapidly decreased over time. Dormant fruit types A and B showed increased germinability up to 7 years, though loss of germinability was lower for type B than for type A fruits. Eight-year-old fruits of all types had significantly lower germinability than younger fruits, probably due to loss of viability. Heterocarpy, therefore, ensures that emergence rates for seedlings of A. sagittata will be maintained over relatively long periods, even in years of strong disturbance when all adult plants may be destroyed. The experiment further showed that germinability of all fruit types in high concentrations of salt, as compared with water treatment, changed over the course of 8 years. Whilst dormant types (A and B) of A. sagittata show increased germinability with age of the seed in water treatment, they significantly lose germinability over time with salinity treatment. Type C fruit was not influenced by salt in the first year, but germinability rapidly decreased with time. It follows that the species is able to germinate under high salt concentration in the first year, but this advantage gradually disappears.     

An extracellular halophilic protease SptA from a halophilic archaeon Natrinema sp. J7: gene cloning, expression and characterization

A gene encoding an extracellular protease, sptA, was cloned from the halophilic archaeon Natrinema sp. J7. It encoded a polypeptide of 565 amino acids containing a putative 49-amino acid signal peptide, a 103-amino acid propeptide, as well as a mature region and C-terminal extension, with a high proportion of acidic amino acid residues. The sptA gene was expressed in Haloferax volcanii WFD11, and the recombinant enzyme could be secreted into the medium as an active mature form. The N-terminal amino acid sequencing and MALDI-TOF mass spectrometry analysis of the purified SptA protease indicated that the 152-amino acid prepropeptide was cleaved and the C-terminal extension was not processed after secretion. The SptA protease was optimally active at 50°C in 2.5 M NaCl at pH 8.0. The NaCl removed enzyme retained 20% of its activity, and 60% of the activity could be restored by reintroducing 2.5 M NaCl into the NaCl removed enzyme. When the twin-arginine motif in the signal peptide of SptA protease was replaced with a twin-lysine motif, the enzyme was not exported from Hfx. volcanii WFD11, suggesting that the SptA protease was a Tat-dependent substrate.Electronic Supplementary Material Supplementary material is available to authorised users in the online version of this article at .     

The isolation, purification and biological activity of a novel antibacterial compound produced by Pseudomonas stutzeri

A novel compound designated zafrin [4beta-methyl-5, 6, 7, 8 tetrahydro-1 (4beta-H)-phenanthrenone] was isolated from a crude extract of a marine bacterium identified as Pseudomonas stutzeri. Zafrin showed strong antibacterial activity against both Gram-positive and Gram-negative bacteria. The compound was purified and its structure was elucidated by spectroscopic methods including 1H-nuclear magnetic resonance (NMR), 13C-NMR, 1D-NMR and 2D-NMR spectroscopy. It could be demonstrated that a purified solution of zafrin was active against several human pathogens, including Staphylococcus aureus, and Salmonella typhi. By contrast, zafrin did not inhibit the growth of eukaryotic organisms Candida albicans and Schizosaccharomyces pombe. The minimal inhibitory concentration for Gram-positive bacteria ranged from 50 to 75 microg mL(-1) and varied between 75 and 125 microg mL(-1) for Gram-negative bacteria. Zafrin lysed Bacillus subtilis cells grown in an osmotically protected medium, suggesting that it does not act upon the cell wall. Further investigation using B. subtilis indicated that the compound is bactericidal and is likely to target the cell membrane.     

The role of glycerol in the nutrition of halophilic archaeal communities: a study of respiratory electron transport

Abstract Respiratory electron transport activity in the Dead Sea and saltern crystallizer ponds, hypersaline environments inhabited by dense communities of halophilic archaea and unicellular green algae of the genus Dunaliella , was assayed by measuring reduction of 2-( p -iodophenyl)-3( p -nitrophenyl)-5-phenyl tetrazolium chloride (INT) to INT-formazan. Typical rates obtained were in the order of 5.5–17.7 nmol INT reduced h ⁻¹ per 10⁶ cells at 35 ° C. In Dead Sea water samples, respiratory activity was stimulated more than two-fold by addition of glycerol, but not by any of the other carbon compounds tested, including sugars, organic acids, and amino acids, or by addition of inorganic nutrients. Stimulation by glycerol had a half-saturation constant of 0.75 μM. A similar respiratory activity was also found when Dead Sea water samples were diluted with distilled water and incubated in the light. As Dunaliella cells did not reduce INT, it is suggested that photosynthetically produced glycerol leaking from the algae is the preferred carbon and energy source for the development of halophilic archaea in hypersaline environments. In samples from saltern crystallizer pond stimulation of INT reduction by glycerol was much less pronounced, probably because the community was less severely carbon-limited.     

Salinomyces polonicus: A moderately halophilic kin of the most extremely halotolerant fungus Hortaea werneckii

A clade where the most halotolerant fungus in the world – Hortaea werneckii, belongs (hereafter referred to as Hortaea werneckii lineage) includes five species: Hortaea werneckii, H. thailandica, Stenella araguata, Eupenidiella venezuelensis, and Magnuscella marina, of which the first species attracts increasing attention of mycologists. The species diversity and phylogenetic relationships within this lineage are weakly known. In this study two moderately halophilic black yeast strains were isolated from brine of graduation tower in Poland. Molecular phylogenetic analyses based on the rDNA ITS1-5.8S-ITS2 (=ITS), rDNA 28S D1–D2 (=LSU), and RNA polymerase II (rpb2) sequences showed that the two strains belong to Hortaea werneckii lineage but cannot be assigned to any described taxa. Accordingly, a new genus and species, Salinomyces and Salinomyces polonicus, are described for this fungus. Furthermore, molecular phylogenetic analyses have revealed that Hortaea thailandica is more closely related to S. polonicus than to H. werneckii. A new combination Salinomyces thailandicus is proposed for this fungus.